Generation of iPSCs from somatic cells offers tremendous potential for therapeutics, the study of disease states, and elucidation of developmental processes (Soldner, F. et al. Cell 136:964-977 (2009); Yamanaka, S. Cell 137:13-17 (2009)). iPSC production techniques introduce active genes that are necessary for pluripotency, or their derivative RNA or protein products, into a somatic cell to induce pluripotent cellular properties that closely resemble those of embryonic stem cells (ESCs) (Takahashi, K. et al., Cell 126:663-676 (2006); Takahashi, K. et al. Cell 131:861-872 (2007); Yu, J. et al. Science 318:1917-1920 (2007); Park, I. et al. Nature 451:141-146 (2008); Yu, J. et al. Science 324:797-801 (2009); Zhao, X. Y. et al. Nature 461:86-90 (2009)). Indeed, iPSCs have been used to produce viable and fertile adult mice, demonstrating their pluripotent potential to form all adult somatic and germline cell types (Zhao, X. Y. et al. Nature 461:86-90 (2009); Boland, M. J. et al. Nature 461:91-94 (2009)).
Fundamentally, the reprogramming process by which a somatic cell acquires pluripotent potential is not a genetic transformation, but an epigenetic one, where the term epigenetic is used to refer to molecular modifications and interactions that impact upon the cellular readout of the genome, such as covalent modifications of DNA and histones, and protein DNA-interactions.
Optimal reprogramming of somatic cells to a pluripotent state requires complete reversion of the somatic epigenome into an ESC-like state, but to date a comprehensive survey of the changes in such epigenetic marks in a variety of independent iPSC lines has not been reported. Therefore, there is a need in the art to understand the epigenomic and methylation characteristics of induced pluripotent stem cells.
Accordingly, Applicants have performed the first whole-genome profiling of the DNA methylomes of multiple ESC, iPSC, and somatic progenitor lines, encompassing reprogramming performed in different laboratories, using different iPSC-inducing technologies, and cells derived from distinct germ layers. This comprehensive base-resolution epigenomic profiling shows that while on a global scale ESC and iPSC methylomes are very similar, iPSC lines display significant reprogramming variability compared to ESCs, including both somatic “memory” and aberrant reprogramming of DNA methylation. Furthermore, all iPSC lines share numerous aberrantly methylated, non-randomly distributed, megabase-scale genic and non-genic regions that Applicants have termed non-CG mega-DMRs. In iPSCs these regions display incomplete or inappropriate reprogramming of the pluripotency-specific non-CG methylation, and are associated with localized differences in CG methylation and transcriptional abnormalities at genes associated with neural development and function.